Several gram-negative bacteria have been isolated with repeated frequency from periodontal pockets, abscesses and from the advancing front of periodontal lesions. When isolated into pure culture and tested in gnotobiotic animal (rat) models, four genera have been shown to cause alveolar bone loss, ultimately leading to loss of connective tissue attachment. These gram-negative bacteria include Capnocytophaga sputigena, Bacteroides asaccharolyticus, Eikenella corrodens, and Actinobacillus actinomycetemcomitans. We will continue our studies on the relationship between bacterial "age" and fatty acid and phospholipid content and distribution, with special emphasis being directed toward the distribution of branched-chain fatty acids and lyso-containing phosphatides. The periplasmic enzyme alkaline phosphatase (APase) will be localized with electron microscopic enzyme histochemistry. In addition, APase will be purified by density gradient centrifugation and PAGE-electrophoresis. The resulting purified enzyme will be utilized in in vitro bone resorption experiments, as well as in macrophage activation studies. The heteropolysaccharides situated on the outer surface of the outer membrane of Capnocytophaga, Eikenella and Bacteroides will be isolated, purified and characterized by chemical and physical methods. These exopolymeric compounds will be used in ongoing fetal rat bone resorptive studies.